tbuckley/cabot-search huggingface dataset:
DIPG-25. MAPPING TUMOR MICROENVIRONMENT USING EMERGING TECHNOLOGIES FOR PEDIATRIC BRAIN CANCERS
Authors: Sandra Laternser, Antonela Petrovic, Augustine Eze, Susanne Dettwiler, Fabiola Prutek, Paola Agata Contini, Denise Morinigo, Miriam Bornhorst, Elisabeth Jane Rushing, Elizabeth McDonough, Christine Surrette, Fiona Ginty, Hanna Minnis, Robyn Gartrell, Sabine Müller, Roger Packer, Javad Nazarian
Journal: Neuro-Oncology (2023)

Abstract Diffuse midline glioma (DMG) is a fatal childhood brain cancer with a survival rate of less than one year from diagnosis. Pharmacological approaches as well as immunotherapy have failed to make a clinical impact. Incomplete understanding of the tumor microenvironment (TME) and tumor associated antigens have contributed to the observed poor prognosis. We hypothesize that characterization of the DMG TME will identify biology-informed targeted treatments. We thus obtained postmortem specimens from 70 patients diagnosed with brain cancers, including 50 DMGs, 20 other types and 10 non-CNS-cancer patients. Up to four anatomical brain locations were selected including the primary tumor, metastatic and adjacent healthy sites. Formalin-fixed-paraffin-embedded (FFPE) specimens were processed for constructing a tissue microarray (TMA). The TMA was stained for a number of markers (H&amp;E, H3K27M, H3K27me3, KI67) and was scored by a neuropathologist. We then used a multiplexed immunofluorescence (MxIF) technology, Cell DIVE™, to iteratively probe 33 biomarkers on a single tissue slide, focusing on immune cell type profiling and activation, and histone mutation status. Analysis of biomarker density and spatial relationships are underway. The main expressed immune markers across all patients were CD163, CD68, and CD8. CD8, a cytotoxic T-cell biomarker, was highly detected in pons, cerebellum, thalamus and the frontal lobe (tumor and healthy) of DMG patients and varied according to clinical intervention, including ONC201 treatment. Furthermore, in comparison to other tumors, DMGs exhibited a higher expression of CD3, T- cell marker, and CD4, a T-helper cell biomarker. Analysis of Iba1, a microglial marker, confirmed a higher difference in microglial activation in primary tumor compared to metastatic and adjacent healthy tissue. In contrast, CD68 was significantly increased in metastatic sites. Analysis is still ongoing. We report establishment of the most comprehensive TMA for pediatric brain tumors, which provides insights into TME comparing critically important clinical variables.
XRT-02. SINGLE-CELL RNA SEQUENCING IDENTIFIES FUNCTIONAL MACROPHAGE SUBSETS THAT ARE ENRICHED IN RESPONSE TO DIFFERENTIAL PHAGOCYTOSIS INDUCTION AGAINST GLIOMA
Authors: Senthilnath Lakshmanachetty, Kent Riemondy, Andrew Donson, Illango Balakrishnan, Sujatha Venkataraman, Rajeev Vibhakar, Nicholas Foreman, Siddhartha Mitra
Journal: Neuro-Oncology (2023)

Abstract Pediatric diffuse midline gliomas (DMGs), are fatal brain tumors of childhood arising in the ventral pons. Currently, radiation therapy (RT) is the mainstay treatment for DIPG. However, RT is not a curative treatment and provides only temporary relief for most patients. Recent advances in immunotherapy have yielded some fantastic opportunities to effectively treat patients with high-grade pediatric brain tumors. In this study, we demonstrate that fractionated RT (4Gy X 3) induces immunogenic cell death and activates multiple damage-associated molecular patterns (DAMPs) on DMG/DIPG cells. Furthermore, combining 4Gy X 3 with anti-CD47 therapy enhances the in vitro phagocytosis of DIPG/DMG cells by peripheral blood mononuclear cell-derived macrophages. Next, using single-cell RNA sequencing (scRNA-seq), we investigated the transcriptomic profile of macrophages that were co-cultured with irradiated or non-irradiated DMG cells in the presence of anti-CD47 monoclonal antibodies for 24 hours. Our findings identified eleven distinct macrophage clusters displaying different gene expression patterns in all the treatment conditions. However, PBS treatment led to a marked increase in macrophages expressing antigen-presentation genes (HLA-DR and HLA-DQB1). 4Gy X 3 treatment led to the enrichment of macrophages expressing genes related to the oxidative phosphorylation pathway, whereas anti-CD47 treatment led to the enrichment of macrophages expressing genes related to interferon-gamma response and ERK-signaling pathway. Lastly, mice intracranially transplanted with DMG/DIPG that received 4Gy X 3 and anti-CD47 therapy showed a significant decrease in tumor growth and an increase in survival rate than those receiving either monotherapy alone. We are currently performing scRNA-seq validation studies on tumors obtained from orthotopic and syngeneic models of DMG. In summary, our results highlight the functional heterogeneity of macrophages and suggest that combining fractionated RT with anti-CD47 therapy has distinct anti-tumor effects and can be used as a therapeutic approach for treating DIPG patients.
LGG-10. TREATMENT-RELATED MYELODYSPLASTIC SYNDROME FOLLOWING CHEMOTHERAPY FOR PEDIATRIC LOW-GRADE GLIOMA
Authors: Phoebe Power, Susannah Payne, Rebecca Walsh, Neevika Manoharan, Adam Nelson
Journal: Neuro-Oncology (2023)

Abstract Treatment-related myeloid neoplasms are rare though well-defined secondary malignancies associated with prior cytotoxic treatment, (particularly alkylating agents and topoisomerase inhibitors). They are rarely associated with primary central nervous system tumours and have historically carried a very poor prognosis. Pediatric low-grade gliomas (pLGG) are the most common CNS tumours of childhood, and up to 50% of patients will require adjuvant therapy. This has traditionally consisted of low-dose metronomic chemotherapy, although the advent of genomic profiling and identification of molecular drivers of pLGG means novel targeted therapies are changing this paradigm. There has only been a single reported case of treatment-related myeloid leukaemia secondary to pLGG treatment. We present a novel case of a 17-year-old girl with treatment-related myelodysplastic syndrome following chemotherapeutic treatment for pLGG. The patient was initially diagnosed at age 4 with a metastatic pilomyxoid astrocytoma (primary suprasellar lesion with spinal metastases) and received first-line treatment with a vincristine and carboplatin regimen. She had disease progression at age 7 and received second-line treatment with thioguanine, procarbazine, lomustine and vincristine. Following further progression at age 16, a biopsy of the suprasellar mass showed a pilocytic astrocytoma with a KIAA1549-BRAF fusion. Prior to commencing a targeted treatment as third-line therapy, she was noted to have a macrocytic anaemia, which subsequently evolved to a bicytopenia. Bone marrow analysis showed a hypocellular marrow with an abnormal myeloid clone with a chromosomal 1q gain, consistent with a diagnosis of treatment-related myelodysplastic syndrome. Germline analysis was negative for bone marrow failure predisposition gene abnormalities. She is currently undergoing workup for bone marrow transplant. Given the poor prognosis of treatment-related myeloid neoplasms, this case represents an important note of caution when choosing appropriate therapy for pLGG, especially given the evolving role for targeted treatments in this disease with otherwise very favourable long-term survival.
ATRT-15. ATRTS HIJACK EPIGENETIC MECHANISMS TO PROMOTE IMMUNOSUPPRESSION AND ESCAPE FROM NK CELL THERAPY
Authors: Jyothishmathi Swaminathan, Ben Ho, Shinji Maegawa, Lei Guo, Alvaro Laureano, Ben Brugmann, Bridget Kennis, Tara Dobson, Shavali Shaik, Visweswaran Ravikumar, David Sandberg, Laurence Cooper, Wafik Zaky, Soumen Khatua, Lucia Silla, Dean Lee, Arvind Rao, Arif Harmanci, Lin Xu, Annie Huang, Vidya Gopalakrishnan
Journal: Neuro-Oncology (2023)

Abstract Atypical Teratoid/Rhabdoid Tumors (ATRTs) are malignant tumors of the central nervous system in children. They are almost universally characterized by genetic loss of SWI/SNF Related, Matrix Associated, Actin Dependent Regulator of Chromatin Subfamily member B1 (SMARCB1) - a core subunit of the ATP-dependent SWI/SNF chromatin remodeling complex, and dysregulation of lineage restricted enhancers. Here, we show that tumor subgroup-specific upregulation of a distant member of the transforming growth factor-β (TGF-β), bone morphogenic factor (BMF) family of proteins drives insensitivity to NK-mediated cytolysis in vitro and in vivo. Mechanistically, subgroup-specific loss of chromatin occupancy by the RE1 Silencing Transcription Factor (REST) to its consensus RE1 motif, allowed deposition of enhancer-specific histone H3 lysine4 (K4) mono-methylation and histone H3K27 acetylation marks to activate an enhancer-promoter interaction. Restoration of REST binding or BET-domain inhibition resulted in a significant reduction in the enhancer marks and enhancer-promoter interaction and involved histone H3K27me3 deposition at the REST binding site, to facilitate gene silencing and sensitization of ATRTs to NK-mediated cytolysis. Thus, our study has identified a novel function for REST as a modulator of tumor-intrinsic enhancer activity and tumor-extrinsic immune microenvironment. Of equal significance, we highlight a role of REST in recruiting the polycomb repressive complex-2 (PRC2) in gene silencing in ATRTs.
DIPG-11. DISSECTING THE DIFFUSE MIDLINE GLIOMA TUMOR MICROENVIRONMENT COMMUNICATIONS USING MULTI-OMICS APPROACHES
Authors: Raphael Collot, Cristian Ruiz Moreno, Amber Wezenaar, Anoek Zomer, Hannah Johnson, Henk Stunnenberg, Anne Rios
Journal: Neuro-Oncology (2023)

Abstract Diffuse midline glioma (DMG) represents a highly aggressive pediatric brain tumor with no chance of survival. Treatment options are urgently needed, and efforts have been put towards developing T-cell immunotherapy. However, this should consider DMG’s unique tumor microenvironment (TME), both in terms of anatomical location and developmental stage. To gain knowledge on this unique tumor niche, we implemented an immuno-competent, syngeneic mouse model based on in-utero electroporation (IUE) of the key driver mutations to induce tumor formation at an embryonic stage. We characterized DMG’s spatial and molecular landscape by combining snRNAseq, spatial transcriptomics, and state-of-the-art imaging. We observed a high intratumoral heterogeneity with spatially restricted cell populations. Importantly, their molecular profiles correlate with patient data, illustrating the clinical relevance of the IUE model. Analysis of cell-cell communications in the TME revealed novel signaling between DMG and immunosuppressive myeloid cells that could impact T-cell response. Using an advanced patient-derived DMG organoid co-culture model with macrophages and engineered T-cells, we are currently investigating the role of this pathway on T-cells mediated killing of DMG organoids. Thus, this project fills a critical need to delineate and overcome the immune-suppressive environment in DMG and potentiate T cell targeting.
DIPG-09. ATOVAQUONE RADIOSENSITISES DIFFUSE MIDLINE GLIOMAS BY INHIBITING MITOCHONDRIAL METABOLISM AND HYPOXIA
Authors: Faiqa Mudassar, Cecilia Chang, Prunella Ing, Sandy Nguyen, Kristina M Cook, Zachary N Warnken, Geraldine O’Neill, Han Shen, Eric Hau
Journal: Neuro-Oncology (2023)

Abstract Diffuse Midline Glioma (DMG) is a uniformly fatal paediatric brainstem tumour with median survival of less than 1 year. Radiotherapy has been the only effective treatment for decades, but most DMGs recur within several months due to radioresistance. The hypoxic tumour microenvironment, a main feature of solid tumours including gliomas, is a major contributor to radioresistance and impedes the efficacy of radiotherapy. Therefore, alleviating tumour hypoxia to enhance the effectiveness of radiotherapy is a therapeutic strategy to improve survival outcomes of DMG patients. Here, our strategy is to decrease the oxygen consumption rate (OCR) of DMG cells by targeting their mitochondria, which in turn will alleviate hypoxia by sparing more oxygen and subsequently improve the radiosensitivity of DMG cells. Specifically, we performed a high-throughput screening to identify potent OCR inhibitors using a library of 1963 FDA-approved drugs. The most promising OCR inhibitor identified was atovaquone, a drug used for treatment of pneumocystis pneumonia and malaria. We found that atovaquone inhibited mitochondrial metabolism of DMG cells by specifically targeting the mitochondrial complex III. It induced the formation of mitochondrial reactive oxygen species suggesting that it increases oxidative stress. It alleviated hypoxia and decreased the expression of hypoxia-inducible factor-1a in several 3-dimensional DMG neurospheres and improved the radiosensitivity of a range of DMG cultures. To overcome the issues of poor bioavailability of commercially available atovaquone resulting in low therapeutically effective brain concentrations, we tested its efficacy against the amorphous solid dispersion (ASD) atovaquone formulation which appears to enhance the atovaquone levels in the brain. We found that both the formulations inhibited OCR and hypoxia at similar doses and improved the radiosensitivity of DMG. With these promising findings, our further work is assessing the in vivo efficacies of commercially available atovaquone and ASD atovaquone formulation using orthotopic DMG models.
MDB-32. TRANSLATION OF NON-CANONICAL OPEN READING FRAMES AS A CANCER CELL SURVIVAL MECHANISM IN CHILDHOOD MEDULLOBLASTOMA
Authors: Damon Hofman, Jorge Ruiz-Orera, Ian Yannuzzi, Rakesh Murugesan, Jip van Dinter, Karl Clauser, Alexandra Condurat, Amy Herman, Vignesh Shanmugam, Kevin Zhou, Jayne Vogelzang, Li Wang, Jennifer Roth, Keith Ligon, Timothy Phoenix, David Root, Todd Golub, Pratiti Bandopadhayay, Sebastiaan van Heesch, John Prensner
Journal: Neuro-Oncology (2023)

Abstract High-risk medulloblastoma is one of the most recalcitrant pediatric cancers, and children with MYC-amplified disease frequently succumb to relapsed disease. Extensive analyses of the coding genome in this disease have characterized additional somatic events in some subsets of patients, though most tumors lack targetable mutations and do not yield insights regarding their aggressive behavior. At the same time, medulloblastoma is known to exhibit extensive rewiring of translational control in MYC-driven tumors, consistent with recent genetic evidence that the impact of this transcription factor on control of mRNA translation may be the most critical aspect of its function during tumorigenesis. Therefore, to propose previously unknown mechanisms for this disease, we have investigated the functional impact of translation of non-canonical open reading frames (ORFs) across medulloblastoma model systems. We demonstrate that these ORFs are commonly translated in medulloblastoma model systems and patient tumors, correlating with disease subtype. Using genome-wide CRISPR/Cas9 screens, we found that ORFs are frequently essential for cell survival in medulloblastoma and describe widespread reliance on upstream open reading frames (uORFs) in particular. From these, we identify a uORF in the ASNSD1 gene that is selectively upregulated and required for maintenance of cell survival by coordinating the function of the prefoldin-like complex, a poorly understood complex implicated in post-translational control. Together, our findings provide a blueprint for oncogenic uORFs as critical disease mediators both in medulloblastoma and, by extension, human cancers more broadly.
DIPG-06. PRECLINICAL EVALUATION OF A NOVEL ACVR1 INHIBITOR IN DIFFUSE INTRINSIC PONTINE GLIOMAS
Authors: Shiva Senthil Kumar, Rebecca Shoffner, Brendan Devine, Rachid Drissi
Journal: Neuro-Oncology (2023)

Abstract Diffuse intrinsic pontine glioma (DIPG) is an incurable brain disease in children. DIPG is refractory to current available therapies. Hence, new therapeutic approaches are urgently needed. About 80% of DIPGs harbor histone 3 lysine to methionine (H3K27M) mutation, in H3-3A (encoding H3.3 histone) and H3C2 (encoding H3.1 histone) genes. One quarter of DIPG tumors harbor somatic mutations in ACVR1 gene, encoding the serine/threonine kinase receptor ALK2. ACVR1 mutations co-segregate with H3.1K27M mutation, representing a specific tumour subtype with distinct clinicopathological and molecular features. These mutations are associated with an aberrant activation of SMAD pathway believed to promote cell proliferation in DIPGs. In the present studies, we evaluated the effectiveness of an orally bioavailable ALK inhibitor, TP-0184 (Tolero Pharmaceuticals) in ACVR1 mutated (ACVR1-mut) and wildtype (ACVR1- WT) DIPG models. Our results indicate that TP-0184 effectively inhibited cell proliferation in primary-patient derived DIPG cell lines. However, we did not observe any selectivity toward ACVR1 mutations. TP-0184 does not induce cell death, but rather cell cycle inhibition, arresting cells in G2/M phase. Interestingly, when treated with ionizing radiation (IR), ACVR1-mut cells preferentially activated SMAD signaling compared to ACVR1-WT. We are currently evaluating the combination efficacy of TP-0184 with IR in our in vitro and in vivo DIPG models. Data on tumor growth kinetics, survival and the effect on SMAD signaling pathway will be presented and discussed.
LGG-06. DEVELOPMENT OF A WEB-BASED MOBILE DEVICE CALCULATOR APPLICATION FOR PREDICTING SHORT- AND LONG-TERM SURVIVAL IN PEDIATRIC OPTIC PATHWAY GLIOMA: A POPULATION-BASED DATABASE ANALYSIS
Authors: Nolan Brown, Cathleen Kuo, Julian Gendreau
Journal: Neuro-Oncology (2023)

Abstract Optic pathway gliomas (OPG) are low-grade lesions arising from the astrocytes of the visual pathway and diencephalon. For most patients, observation appears to be the treatment of choice, but it remains unclear how frequently serial imaging should be obtained and at what point lesions can be deemed to have become quiescent. There is therefore an ongoing need for prognostication tools that can accurately predict the behavior of OPG to help identify those lesions meriting therapeutic intervention. In the present study, we leverage population-level data from the Surveillance, Epidemiology, and End Results program (SEER) database to develop a predictive calculator for overall survival (OS) in pediatric OPG. Accordingly, we extracted all available patient-level data from the SEER database. Training and testing datasets were established. Cox proportional hazards regressions were then used to identify independent predictors of survival, and model validation was performed. A total of 1,052 pediatric patients met inclusion criteria. Most were between 5 to 9 years old (45.7%), males (51.6%), white (85.6%), non-Hispanic (82.1%), and 232 patients (22.1%) came from houses with a median income less than $54,999, while 111 patients (10.6%) lived in a rural county. Nearly 90% of patients did not undergo surgical debulking. Twenty-three patients (2.2%) received radiotherapy and 294 (27.9%) received chemotherapy. Additionally, univariate and multivariate analyses were performed to identify variables associated with OS. Median household income and subtotal resection were found to be predictors of OS on univariate analysis. Furthermore, on multivariate analysis, median household income greater than $55,000 and subtotal resection were predictive of higher OS. Age at diagnosis, sex, rural versus urban setting, benign versus malignant behavior, and use of chemotherapy or radiotherapy did not appear to affect OS. Finally, we created a calculator (https://spine.shinyapps.io/optic_glioma/) using our model to estimate 5-,10-, and 1-year OS probability and other survival outcomes.
ATRT-16. THE PI3K INHIBITOR PAXALISIB COMBINES SYNERGISTICALLY WITH THE MEK INHIBITOR MIRDAMETINIB TO TARGET ATYPICAL TERATOID/RHABDOID TUMORS
Authors: Kristen Malebranche, Tyler Findlay, Anupa Geethadevi, Charles Eberhart, Eric Raabe, Jeffrey Rubens
Journal: Neuro-Oncology (2023)

Abstract Atypical teratoid/rhabdoid tumors (AT/RT) are aggressive pediatric brain tumors and the most common malignant brain tumors of infancy. The four-year event-free survival rate is only 37%. There are limited treatment options for patients with relapsed AT/RT, but precision therapies may help improve survival for patients with this deadly disease. We have previously identified strong activation of the PI3K-AKT-mTOR (mTOR) signaling pathways in AT/RT. Paxalisib is a highly brain-penetrant PI3K inhibitor acting upstream of mTOR. We find that Paxalisib slows tumor growth in orthotopic xenograft models of AT/RT and extends median survival from 40 to 54 days (p=0.001, log-rank test). RNASeq after mTOR pathway inhibition identifies reflexive activation of the RAS-RAF-MEK-ERK (MAPK) pathway as a possible mechanism of therapy resistance (KEGG pathway analysis). The RAS-RAF-MEK-ERK (MAPK) pathway plays a pivotal role in regulating cell growth and survival and activation of the pathway which contributes to the aggressive growth of numerous cancers. Mirdametinib is a small, allosteric MEK inhibitor currently used in clinical trials for both pediatric low- and high-grade gliomas (PD0325901). We find that dual inhibition of both the mTOR and MAPK pathways by Paxalisib and Mirdametinib synergize to reduce AT/RT growth and viability (Bliss synergy score 16.77). Paxalisib also combines with Mirdametinib to induce high levels of apoptosis and cell senescence (as determined by western blot: cPARP, pRB, P21, P16). Naïve treatments and a current survival study in mice bearing AT/RT orthotopic tumors suggest that combination therapy decreases mTOR and MAPK pathway activation (as determined by western blot). These data support using Paxalisib and Mirdametinib combination therapy as a promising novel therapy to treat relapsed AT/RT.
HGG-12. INFANT HEMISPHERIC GLIOMA: CLINICAL AND MOLECULAR CHARACTERISTICS AND RESULTS OF TARGETED THERAPY
Authors: Ludmila Papusha, Alexander Druy, Margarita Zaytseva, Ekaterina Salnikova, Agnesa Panferova, Alexey Samarin, Galina Novichkova, Eugene Hwang, Roger Packer
Journal: Neuro-Oncology (2023)

Abstract We analyzed an infant cohort consisting of 14 patientswith hemispheric gliomas (hHGG, n=10 and DIGG, n=4) diagnosed at D. Rogachev Center between 2017and 2022. The hHGG cohort included nine females and one male ranging in age from one week to 22 months. RTK-fusions were detected in all (ROS1 - 4, ALK - 2,NTRK1 – 2, NTRK3 - 1, and EGFR - 1). In one of thesecases, the diagnosis was made retrospectively in a patient initially diagnosed with anaplastic ependymoma who died during treatment. Out of the nine remaining hHGG patients, four (ROS1 - 3, NTRK3-1) experienced rapid progressive disease (PD) duringconventional chemotherapy and one pt (ROS1) after RT. These five pts received targeted therapy with entrectinib with initial good responses in all cases (CR-2, PR-3); one of these patients (ROS1) experienced PD after 7 months of targeted therapy and died of disease. Molecular genetic analysis after the second operationin this case revealed an acquired ROS1 G2032R mutation. Treatment-related adverse events on entrectinib therapy were detected in two patients and included bone fractures and neutrophil count decrease in one case and decrease left ventricular ejection fraction in another patient. Median duration of targeted therapy was 7 months (range, 3-33). From those pts who received only chemotherapy – 2 pts are alive with CR and 1 pt died from complication of CT. All patients with DIGG are alive without signs of progression after surgery alone. Median follow-up time is 19 months (range, 5-64). Molecular genetic analysis revealed MAPK-pathway alterations in each case (rearrangements of NTRK1-1, ROS-1, RAF1-1, BRAF-1), respectively. Patients with infant hHGG demonstrate an aggressive course on conventional therapy, especially in cases with ROS1-positive tumors. Targeted therapy may be an effective treatment option but required close monitoring for toxicity and resistance.